The insulin-like growth factors (IGFs), IGF-I and IGF-II, are mitogenic peptide growth factors involved in a broad range of cellular processes including hyperplasia, DNA synthesis, differentiation, cell cycle progression and inhibition of apoptosis (Keiss et al., 1994, Hormone Research 41 66; Wood & Yee, 2000, J. Mammary Gland Biology and Neoplasia 5 1; Jones & Clemmons, 1995, Endocrine Rev. 16 3). These effects are mediated through binding to their tyrosine-kinase linked cell surface receptor, the type 1 IGF receptor (IGF-IR). The IGFs are also tightly regulated by a family of specific binding proteins, termed IGFBPs, whose primary role is to bind free IGFs and thereby moderate their half-life, specificity and activity (Clemmons, 1998, Mol. Cell. Endocrinol. 140 19).
Recently, vitronectin (VN) has been shown to bind directly to IGF-II (Upton et al., 1999. Endocrinology 140 2928-31) while IGF-I can bind to VN in the presence of certain IGFBPs, as described in International Publication WO 02/24219. The finding that VN, an ECM organization and adhesion molecule, binds IGF-II with an affinity that is similar to that of IGF-II for IGF-IR (Upton et al., 1999, supra), its biologically relevant receptor, reveals a specific physical link between IGF action and VN in the ECM. In addition, IGF-II bound to VN can stimulate synergistic functional responses in human keratinocytes in vitro (International Publication WO 02/24219).
VN is a glycoprotein that is highly abundant in the blood and in the ECM. Primarily synthesized in the liver, but expressed by many other cell types, VN circulates in the blood in a closed conformation and is deposited in the ECM in an open, or extended, conformation (Schvartz et al., 1999, The International Journal of Biochemistry and Cell Biology 31 531-44). Both conformations are believed to bind IGF-II (Upton et al., 1999, supra; International Publication WO 02/24219; McMurty et al., 1996, Endocrinology 150:149-60) and also bind multiple other ligands including collagen (Morris et al., 1994, Journal of Biological Chemistry 269 23845-52), glycosaminoglycans (Francois et al., 1999, Journal of Biological Chemistry 274: 37611-19), many other ECM proteins and a wide variety of integrins, particularly the αv integrins. Indeed, the primary role of vitronectin is as an ECM organization molecule that provides adhesive links to these cell surface integrin receptors via an RGD binding motif. The VN receptors (αv integrins) have been shown to regulate the actin cytoskeleton rearrangement required for growth and invasion, hence, VN binding coordinates cell adhesion and movement (DePasquale, 1998, Histochemistry and Cell Biology 110: 485-94; Huang, 2000, Oncogene 19 1915-23).
However, the respective, relative contributions of IGFs and VN present in protein complexes, in terms of stimulating biological responses such as cell migration and/or proliferation, has remained elusive, as has the site of protein-protein interaction between IGFs/IGFBPs and VN.